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Characterization of 40 soybean (Glycine max) promoters, isolated from across 5 thematic gene groups

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Abstract

Gene expression in plants is primarily modulated by the promoter region, which is located upstream of the gene coding region. Specific regulatory sequences within the promoter region contribute to the strength, tissue specificity, timing and duration of gene expression. In this study, promoter regions were isolated from 40 different soybean (Glycine max) genes that were grouped into five distinct categories, based on the localization of native gene expression (constitutively expressed genes, root-preferentially expressed genes, seed-preferentially expressed genes), or the associated biological processes of native gene expression (fatty-acid biosynthesis pathway associated genes, and WRKY transcription factor genes). Each of the promoters was placed upstream of the green fluorescent protein coding sequence and promoter functionality was then assessed using transient expression analysis following particle bombardment of lima bean (Phaseolus lunatus) cotyledons, as well as stable expression analysis following Agrobacterium rhizogenes induction of soybean hairy roots. Surprisingly, promoter strength was relatively consistent using the two different validation methods while tissue specific expression was not observed, likely due to a combination of the target tissue, the large amount of DNA that was introduced and minimal epigenetic effects. The differences in gene expression within members of each promoter group indicates that promoters need to be characterized in a transgenic environment, rather than relying on expression predicted using native gene expression profiles. This systematic approach into the characterization of native soybean promoters adds to the knowledge base of soybean gene regulation and the toolbox of promoters for plant biotechnology.

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Acknowledgments

The authors are grateful for the work of Dr. Robert A. Bouchard in identifying the promoters belonging to GmSeed promoter family, as well as for carrying out a number of the experiments. The authors also thank Ning Zhang for valuable technical inputs. Salaries and research support for this work were provided by the United Soybean Board, and State and Federal Funds appropriated to the Ohio Agricultural Research and Development Center, The Ohio State University. Mention of trademark or proprietary products does not constitute a guarantee or warranty of the product by OSU/OARDC and also does not imply approval to the exclusion of other products that may also be suitable.

Author contributions

J.J.F. and P.J.R. conceived and designed the research. A.G. gathered data and performed the majority of the analyses. L.K.M. and A.H.G. analyzed RNAseq data. J.J.F., P.J.R., and L.K.M. defined the promoter groups. A.G., P.J.R., L.K.M. and J.J.F performed data interpretation. A.G., A.H.G., P.J.R., L.K.M. and J.J.F. wrote the manuscript. All authors read and approved the manuscript.

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Authors and Affiliations

  1. Department of Horticulture and Crop Science, The Ohio State University, 1680 Madison Ave., Wooster, OH, 44691, USA

    Andika Gunadi & John J. Finer

  2. 22nd Century Group, 9530 Main St., Clarence, NY, 14031, USA

    Paul J. Rushton

  3. Department of Horticulture and Crop Science, The Ohio State University, 2021 Coffey Rd, Columbus, OH, 43210, USA

    Leah K. McHale & Amanda H. Gutek

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  1. Andika Gunadi
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Correspondence to John J. Finer.

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Electronic supplementary material

Below is the link to the electronic supplementary material.

Online Resource 1

Primers used for promoter amplification (XLSX 12 kb)

Online Resource 2

Promoter sequences in FASTA format (PDF 157 kb)

Online Resource 3

Occurrence of putative cis-regulatory elements identified using PLACE in GmCons promoters (XLSX 54 kb)

Online Resource 4

Occurrence of putative cis-regulatory elements identified using PLACE in GmRoot promoters (XLSX 111 kb)

Online Resource 5

Occurrence of putative cis-regulatory elements identified using PLACE in GmSeed promoters 5 (XLSX 133 kb)

Online Resource 6

Occurrence of putative cis-regulatory elements identified using PLACE in GmFAB promoters (XLSX 141 kb)

Online Resource 7

Occurrence of putative cis-regulatory elements identified using PLACE in GmWRKY promoters (XLSX 168 kb)

Online Resource 8

Animations showing transient GFP expression in lima bean cotyledonary tissue over 100 hours for all promoters (ZIP 523853 kb)

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Gunadi, A., Rushton, P.J., McHale, L.K. et al. Characterization of 40 soybean (Glycine max) promoters, isolated from across 5 thematic gene groups. Plant Cell Tiss Organ Cult 127, 145–160 (2016). https://doi.org/10.1007/s11240-016-1038-x

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